Aging of the skin is a complex process induced by chronological and environmental factors (mainly UV radiation). Signs or symptoms of skin aging include the loss of skin elasticity and firmness, the appearance of features such as wrinkles and furrows, dark under-eye circles, puffy eyes, eye bags, solar lentigines (age spots) and mottled skin. The first signs of skin aging are usually evident on a person's face, specifically in the region around the eyes. These include the presence of dark eye circles (periorbital hyperpigmentation), puffy eyes (periorbital puffiness), eye bags (infraorbital palpebral bags), and wrinkles (for example, periorbital wrinkles). The presence of the signs of aging on a person's skin, especially their face, is aesthetically undesirable. Younger looking skin, that is, skin with reduced symptoms of aging, is desired.
The skin, mucous membranes, hair and/or the nails provide a physical barrier between an organism and its environment. The skin is composed of two principal layers, the epidermis and the dermis. The dermis is the thickest layer (having an approximate thickness of 90% of the thickness of the skin) and contains collagen, elastin, several differentiated structures such as blood vessels and many cell types such as fibroblasts (which synthesize collagen and elastin). The epidermis is composed of keratinocytes, melanocytes and Langerhans cells, with the main cell population composed of keratinocytes.
Collagen is the most abundant protein in the skin's connective tissue and plays an important structural role in the skin. It forms a mesh like structure in the skin connective tissue that helps support new cells as they grow while providing the needed flexibility. There is continual collagen synthesis and degradation in the skin, and the balance between them determines both the tensile strength and elasticity of the skin. Elastin is a protein in the connective tissue that is elastic. Elastin is helps to keep the skin flexible but tight, providing a bounce-back reaction if the skin is pulled. The aging process is accompanied by degeneration and lysis of both collagen fibers and elastic fibers in the skin. The gradual disappearance of elastic fibers in the skin results in the progressive loss of skin elasticity. The degeneration and lysis of collagen fibers results in the skin losing resistance (firmness). A further consequence of connective fibers lysis in the dermis is the gradual reduction of the dermis thickness as a whole, particularly through the reduction in collagen fibers. [Bonta M, Daina L, Muiu G. The process of ageing reflected by histological changes in the skin. Rom J Morphol. Embryol. 2013; 54 (3 Suppl.):797-804]
Another factor in the aging of the skin is the appearance of Advanced Glycosylation End Products (AGEs). AGEs are obtained from a reaction called glycation involving sugar and protein. The presence of these products in the skin changes the physical, biomechanical (the skin stiffens and loses elasticity) and biological properties (modulation of the synthesis, degradation of the matrix by cells). AGEs can modulate the expression of proteins of the extracellular matrix (ECM) as collagen, and they can also modify the expression and synthesis of the enzymes which are responsible for its degradation (elastase and metalloproteinases enzymes) [Pageon, H. Reaction of glycation and human skin: the effects on the skin and its components, reconstructed skin as a model. Pathol. Biol. (Paris). 2010 June; 58(3):226-31]. The result is reduced elasticity and thickness of the skin. In skin, glycation of collagen Type I has been linked to the development of skin dullness and the decrease of skin elasticity.
As a result of reduced elasticity, firmness and thickness, wrinkles in the skin can appear, such as those that appear around the eye. There is a need to provide an active agent which can help prevent collagen degradation and/or stimulate collagen production in the skin. There is a need to provide an active agent which can help prevent elastin degradation and/or stimulate elastin production in the skin. There is a need to provide an active agent that can inhibit the formation of AGEs in the skin. Such active agents can be useful in the treatment skin to prevent or alleviate signs of aging.
The aging process also affects the vasculature in the skin. Vascular changes include the thinning of capillary walls and the slowing of microcirculation. Alteration of the blood vessel walls causes changes in vascular permeability and can result in the appearance of interfibrillar edema [Bonta M, Daina L, Muiu G. The process of ageing reflected by histological changes in the skin. Rom J Morphol. Embryol. 2013; 54(3 Suppl.):797-804]. Thus one of the signs of aging is the accumulation of interstitial fluid around and under the eyes, for example, puffy eyes and eye bags (also known as bags under the eye). These are aesthetically unsightly and it is desired that the puffiness of the skin/volume of the bags is reduced. There is a need for active agents that are able to decrease vascular permeability implicated in the edema formed in puffy eyes and eye bags.
As skin ages, it becomes thinner. For example, Bonta et al. note that decreased vascular efficiency, especially in the superficial dermis, produces a series of major effects in the epidermis, by adapting it to the efficiency of vasculature, namely by reducing the number of cell layers, i.e. by reducing the thickness [Bonta M, Daina L, Muiu G. The process of ageing reflected by histological changes in the skin. Rom J Morphol. Embryol. 2013; 54(3 Suppl.):797-804]. This thinness can result in the underlying blood vessels and chromophores (such as bilirubin and melanin) becoming more visible. This is one cause of dark under-eye circles, a very common cosmetic problem affecting the majority of people [Ranu H, Thng S, Goh B K, Burger A, Goh C L. Periorbital hyperpigmentation in Asians: an epidemiologic study and a proposed classification. Dermatol. Surg. 2011 September; 37(9):1297-303]. It is believed that this problem is further exacerbated, by the blood vessels becoming leaky with aging and, as a result, bilirubin, a breakdown product of blood, accumulating around the eyes. Specifically, bilirubin is a breakdown product of heme metabolism. Heme is an iron-containing porphyrin found in hemoglobin, myoglobin, and several enzymes of which the hepatic cytochromes are the most important representatives. Approximately 80% of daily bilirubin production derives from senescent red blood cells. These are broken down and iron is removed from the heme molecule and the remaining porphyrin ring is oxidized and cleaved at a single site to form the tetrapyrrole chain structure of biliverdin. Further reduction of the biliverdin results in the formation of bilirubin responsible for the coloration appearing in the infraorbital eyelids as dark under-eye circle [Stillman A E. Jaundice. Clinical Methods: The History, Physical, and Laboratory Examinations. 3rd edition. Boston: Butterworths; 1990. Chapter 87]. Dark eye circle is a complex facial cosmetic problem, with multiple causes and these include melanin deposition, venous stasis with hemosiderin deposition, and orbital structural problems. Melanin deposits in the dermis may be congenital or secondary to environmental factors such as excessive exposure to the sun, endogenous or use of exogenous estrogens, pregnancy and breastfeeding. There is a need to provide an active agent that can degrade bilirubin and/or reduce the amount of melanin in the skin, two of the pigments responsible of the pigmentation in dark circles around the eyes.
The signs of aging such as wrinkles, dark under-eye circles, puffy eyes, eye bags can be exacerbated by fatigue, stress, drug and alcohol use, among other factors.
The modification of skin color, including the lightening of skin, for example, of dark eye circles as well as the elimination or attenuation of age spots, is a cosmetic effect desired by many people. Often the aim is to achieve an even skin color. Depigmenting cosmetic products are used to reduce hyperchromia and, typically, depigmenting agents act by inhibiting a melanin biosynthesis route.
Melanins are complex pigments that provide the skin, hair and eyes of mammals with color and photoprotection against ionizing radiation. Melanogenesis is physiological process resulting of the synthesis of the melanin pigments, and is characterized, in summary, by the production process and subsequent distribution of melanin by melanocytes. Mammalian melanocytes produce two chemically distinct types of melanin pigments, the black to brown eumelanin and the yellow to reddish brown pheomelanin, by different enzymes in complex organelles called melanosomes. Both eumelanin and pheomelanin are derived from the common precursor dopaquinone that is formed by tyrosinase (TYR). Tyrosinase is also called polyphenol oxidase and is a copper-containing multifunctional enzyme. It is the key enzyme in the first stage of melanogenesis cascade, catalyzing the conversion of L-tyrosine in L-dopaquinone (Ito S., Wakamatsu K., and Ozeki, H. Chemical analysis of melanins and its application to the study of the regulation of melanogenesis. Pigment Cell Res. 2000: 13 Suppl. 8. 103-9). In addition to tyrosinase, two related proteins, termed tyrosinase-related proteins (TRPs), have been shown to regulate eumelanin formation. TRP-1 (Tyrosinase-related protein-1) or DHICA (5,6-dihydroxyindol-2-carboxylic acid oxidase), and TRP-2 (Tyrosinase-related protein-2), also known as dopachrome tautomerase (Hearing. V. J. The melanosome: the perfect model for cellular response to the environment Pigment Cell Res. 2000; 13 Suppl. 8, 23-4). Melanin formation also originates in tyrosine oxidation, by the enzyme tyrosinase, to dihydroxyphenylalanine (DOPA) inside melanocytes.
Melanosomes are lysosome-related organelles which have the unique capacity to produce melanin pigment and which progress through four sequential morphological steps as they mature (stage I, II, III and IV). Stage I melanosomes are round, membrane-bound and electron-lucent vesicles that are generally found in the perinuclear area. The transition to Stage II melanosomes involves an elongation of the vesicle, and the appearance within of distinct fibrillar structures. The production of those internal matrix fibers and the maturation from Stages Ito II melanosomes depend on the presence of a structural protein termed Pmel17, (also known as gp100 or SILV). Shortly after its delivery to Stage I melanosomes, Pmel17 is cleaved into several fragments, which form the fibrillar matrix of the organelle. In pigmented cells, melanin is deposited on these fibers, resulting in a progressively pigmented internal matrix, at which time the organelles are termed Stage III melanosomes. In highly pigmented tissues, melanin synthesis and deposition continues until little or no internal structure is visible, at which time they are termed Stage IV melanosomes. Several proteins have been identified as melanosome-specific proteins (Tyr, Trp1, Trp2, MART-1, Pmel17, GPNMB, etc.). MART-1, melanoma-associated antigen recognized by T cells protein (also known as Melan-A), a melanosome-specific proteins, has no detectable enzymatic activity, is highly enriched in early melanosomes (Stage I and/or II melanosomes), and forms a complex with Pmel17 and affects its expression, stability, trafficking, and the processing which is required for melanosome structure and maturation and thus plays an important role in regulating mammalian pigmentation (Hoashi T, Watabe H, Muller J, Yamaguchi Y, Vieira W D, Hearing V J. MART-1 is required for the function of the melanosomal matrix protein PMEL17/GP100 and the maturation of melanosomes. J Biol. Chem. 2005 Apr. 8; 280(14):14006-16. Epub 2005 Jan. 28.) GPNMB (glycoprotein (transmembrane) nonmetastatic melanoma protein b), a highly glycosylated type I transmembrane protein, exhibits a high similarity with Pmel17. GPNMB contains several domains related with its functions in melanocytes. It has been confirmed that the arginine-glycine-aspartate (RGD) motif of GPNMB can bind to integrins to regulate the adhesion of melanocytes with keratinocytes, indicating that it is involved in the transfer of melanin. As an important structural protein of melanosomes, GPNMB has proven to be present in all stages (I-IV) of melanosomes, and is especially enriched in mature stages. GPNMB deletion in melanocytes sharply attenuated melanosome formation, indicating a critical role in melanosome synthesis (Zhang P, Liu W, Zhu C, Yuan X, Li D, Gu W, Ma H, Xie X, Gao T. Silencing of GPNMB by siRNA inhibits the formation of melanosomes in melanocytes in a MITF-independent fashion. PLoS One. 2012; 7(8):e42955.)
After the production, melanin, within the melanosomes, is transferred to the adjacent keratinocytes through dendrites present in the melanocytes, where it shall be transported and degraded. This melanin transference may occur through three different mechanisms: Cytophagocytosis process of the dendritic end of the melanocyte by the keratinocyte; direct migration of melanosomes of the cytoplasm to the keratinocyte and; release of the melanosomes in the extracellular space and its incorporation to the keratinocytes. Thus, skin pigmentation depends on the number, the chemical nature of melanin and content (the tyrosinase activity), and distribution of melanosomes produced, and transferred by each melanocyte to a cluster of keratinocytes surrounding it.
Increased melanin production due to the direct or indirect stimulation is a defensive reaction of skin in order to protect against solar aggression. After UV irradiation, the melanosomes regroup around the nucleus in order to protect the cell's genetic material and thus, in addition to promoting the skin and hair coloring, melanin promotes photo protection, acting as a sun filter, diffracting or reflecting solar radiation. The melanocyte-keratinocyte complex responds quickly to a wide range of environmental stimuli, often in paracrine and/or autocrine manners. Thus, melanocytes respond to UV-R, agouti signaling protein, melanocyte-stimulating hormone (MSH), endothelins, growth factors, cytokines, etc. After UV-R exposure, melanocytes increase their expression of proopiomelanocortin (POMC, the precursor of MSH) and its receptor melanocortin 1 receptor (MC1-R), TYR and TYRP1, protein kinase C (PKC), and other signaling factors. On the other hand, it is known that UV stimulates the production of endothelin-1 (ET-1) and POMC by keratinocytes and that those factors can then act in a paracrine manner to stimulate melanocyte function. In addition to keratinocytes, fibroblasts, and possibly other cells in the skin produce cytokines, growth factors, and inflammatory mediators that can increase melanin production and/or stimulate melanin transfer to keratinocytes by melanocytes. Melanocyte growth factors affect not only the growth and pigmentation of melanocytes but also their shape, dendricity, adhesion to matrix proteins, and mobility.
α-MSH, ACTH, basic fibroblast growth factor (bFGF), nerve growth factor (NGF), endothelins, granulocyte-macrophage colony-stimulating factor (GM-CSF), steel factor, leukemia inhibitory factor (LIF), and hepatocyte growth factor (HGF) are keratinocyte-derived factors that are thought to be involved in the regulation of the proliferation and/or differentiation of melanocytes, some acting through receptor mediated signaling pathways. It has been shown that in human epidermis, α-MSH and ACTH are produced in and released by keratinocytes and are involved in regulating melanogenesis and/or melanocyte dendrite formation. α-MSH and ACTH bind to a melanocyte-specific receptor, MC1-R, which activates adenylate cyclase through G protein, which then elevates cAMP from adenosine triphosphate. Cyclic AMP exerts its effect in part through protein kinase A (PKA), which phosphorylates and activates the cAMP response element binding protein (CREB) that binds to the cAMP response element (CRE) present in the M promoter of the microphthalmia-associated transcription factor (MITF) gene. The increase in MITF-M expression induces the up-regulation of TYR, TYRP1, and DCT, which leads to melanin synthesis.
Hyperpigmentation is a disorder caused by exaggerated melanin production. Factors such as excessive solar exposure, aging, hormone changes, inflammation, allergies, among others, may cause an unbalance in the melanin production and distribution process, resulting in skin stains. Solar lentigines (also known as senile lentigo, sun-, liver-, or age spots) are circumscribed, pigmented macules, which are usually light brown, but vary in degree of color to jet black. Solar lentigines are typically found on UV-exposed areas of the body (the face, dorsum of the hand, extensor forearm, upper back, and decolletage). They can range anywhere in size from 1 mm up to a few centimeters in diameter and, in areas of severely sun-damaged skin, may coalesce into even larger lesions. There is a desire to provide cosmetic actives that can lighten the color of hyperpigmented skin such as solar lentigines (age spots).
The molecular mechanism currently proposed for the appearance of solar lentigines involves the stimulation of two epidermal cascades, consisting of ET-1/ETBR (initiated by the binding of ET-1 to its receptor, ETBR), and SCF/c-kit (initiated by the binding of Stem cell factor to its receptor c-Kit), and the cross-talk between those two after the UV exposure. Exposure to UV radiation induces an increase in the production of ET-1 by keratinocytes, and its secretion therefore stimulates melanocytes to produce melanin. The potential of keratinocytes located in SL lesional epidermis to produce ET-1 is significantly higher than in perilesional normal controls, and there is an accentuated expression of ETBR transcripts as well. The increased production and localization of ET-1 was paralleled by increased amounts of tyrosinase in melanocytes. Separately, SCF (also produced by keratinocytes) binds to the c-KIT receptor on melanocytes. Solar lentigines lesional epidermis expresses increased levels of SCF mRNA transcripts and protein compared with nonlesional controls. [Costin G E, Hearing V J. Human skin pigmentation: melanocytes modulate skin color in response to stress. FASEB J. 2007 April; 21(4):976-94]
However, other cascades in the skin may also contribute to the hyperpigmentation seen in solar lentigines. The Wnt signaling pathway, which triggers the differentiation of melanocyte stem cells, in solar lentigine lesions, is implicated in the accelerated differentiation of melanocyte stem cells was involved in the formation of SLs. The Wnt signaling pathway is closely related to melanocyte biology. This signaling pathway is also important for melanocyte stem cells to trigger the differentiation into follicular melanocytes and epidermal melanocytes. The protein dickkopf WNT signaling pathway inhibitor 1 (DKK1), an inhibitor of the Wnt signaling pathway, prevented melanogenesis and decreased the density of melanocytes. DKK1 suppresses melanocyte function and growth through the regulation of microphthalmia-associated transcription factor (MITF) and b-catenin. [Yamaguchi Y, Morita A, Maeda A, Hearing V J. Regulation of skin pigmentation and thickness by Dickkopf 1 (DKK1). J Investig. Dermatol. Symp. Proc. 2009 August; 14(1):73-5.] [Yamada T, Hasegawa S, Inoue Y, Date Y, Arima M, Yagami A, Iwata Y, Takahashi M, Yamamoto N, Mizutani H, Nakata S, Matsunaga K, Akamatsu H. Accelerated differentiation of melanocyte stem cells contributes to the formation of hyperpigmented maculae. Exp. Dermatol. 2014 September; 23(9):652-8.]
Thus an active agent which can inhibit a melanin biosynthesis route or act directly on it can have a lightening or depigmenting effect on the skin, such as an active agent that can inhibit tyrosinase activity, inhibit the production of melanin in melanocytes, or affect the expression of genes involved in the melanogenic process, can act as a depigmenting agent. There is a need for a new active agent effective in having a lightening, whitening or depigmenting effect on the skin
The present invention sets out to solve some or all of the above-identified problems and meet some or all of the above-identified needs.